Recent advances in molecular virology have enabled investigators to construct viruses that selectively destroy cancer cells (oncolytic virotherapy). Genetically engineered viruses belonging to different viral families have been evaluated for their potential as therapeutic agents in the treatment of malignant tumors. Efficient replication, cell lysis, and spread of herpes simplex viruses (HSV), and their natural broad host range make them attractive candidates as oncolytic viral agents. Furthermore, the recent availability of cloned HSV genomes into bacterial artificial chromosome vectors greatly facilitates the rapid construction of new recombinant viruses carrying multiple transgenes of interest (Jeyaretna and Kuroda, 2007. Tumor treatment with oncolytic HSV has been shown to induce anti-tumor immune responses (Li et al., 2007; Miller and Fraser, 2003; Nakamori et al., 2004; Thomas and Fraser, 2003; Todo et al., 1999). Although the majority of people are seropositive for HSV-1, oncolytic virotherapy with HSV is not limited by pre-existing anti-HSV immunity (Hoffmann, Bayer, and Wildner, 2007; Kemeny et al., 2006), and in at least one example, preexisting immunity to HSV-1 enhanced anti-tumor immune responses (Miller and Fraser, 2000).
Recently, the NV1020 oncolytic herpes simplex virus type-1 (HSV-1) was shown to have significant promise for the treatment of many different types of tumors in preclinical studies in experimental animals as well as in human clinical trials (Cozzi et al., 2002). The main advantage of this virus over other HSV oncolytic viruses is that it expresses one of the two original γ134.5 genes allowing the virus to replicate more efficiently, while safety is not compromised. The γ134.5 gene is a major neurovirulence gene and an inhibitor of cellular apoptosis. Deletion of this gene drastically attenuates the virus and restricts viral growth to cancer cells because of their lack of intact apoptotic mechanisms. Preclinical studies in mice as well as phase I/II human trials have revealed that oncolytic HSV-1 viruses having both γ134.5 genes deleted did not spread efficiently within tumors (Kramm et al., 1997). In contrast, deletion of one of the two γ134.5 genes drastically attenuated the virus, while allowing improved virus replication and spread in tumor cells (Advani et al., 1999; Meignier, Longnecker, and Roizman, 1988; Meignier et al., 1990). The HSV-1 virus, NV1020, was originally constructed for vaccine purposes and contained HSV-2 viral sequences coding for glycoproteins gD, gG, gI and gE to facilitate production of anti-HSV-2 immune responses, as well as other genetic alternations (Meignier, Longnecker, and Roizman, 1988). A fusogenic oncolytic HSV-1 Synco-2D was reported to elicit anti-tumor immune responses when studied in a similar animal model of mammary carcinoma utilizing 4T1 cells (Nakamori et al., 2004). A strong T-cell response was reported also by an HSV-2 derivative oncolytic virus FusOn-H2 effectively treating primary and metastatic mammary tumors in vivo (Li et al., 2007). A number of different HSV-1 recombinant viruses have been constructed and evaluated for their ability to treat a variety of different cancers in animal models, as well as in human phase I/II clinical trials (Todo, 2002; Hu and Coffin, 2003; Argnani et al., 2005; Shen and Nemunaitis, 2006). The most important modification, which is common to all constructed HSV-1 viruses is the modification/deletion of the γ1 34.5 gene, based on the knowledge that γ1 34.5 is an important determinant of neurovirulence and an inhibitor of cellular apoptosis.
Although γ1 34.5 is non-essential for virus replication, deletion of both copies of the γ1 34.5 genes causes substantial reduction in infectious virus production, because the γ1 34.5 protein is a structural component of the virion particle and is involved in intracellular glycoprotein transport and cell-surface expression requiring a minimum amount of the protein to be expressed for optimum infectious virus production (Andreansky et al., 1997; Kramm et al., 1997; Todo et al., 2001). In a comparison of the previously constructed R3616 and R7020 that have either both or one of the two γ1 34.5 deleted, respectively, R7020 replicated to much higher levels compared to the double-deletion mutant R3616 both in vitro and in vivo (Advani et al., 1999). Furthermore, R7020 replicated preferentially in neoplastic cells and exhibited a remarkable safety profile in extensive rodent and primate studies as well as in human vaccine trials (Meignier et al., 1988; Meignier et al., 1990). R7020 is being currently evaluated in human clinical trials under the name NV1020. Recently, it was shown that NV1020 could be safely administered into the hepatic artery without significant effects on normal liver function in a phase I, open-label, dose-escalating study with subjects having metastatic colorectal carcinoma to the liver (Kemeny et al., 2006). The G207 virus carries a double-deletion of the γ1 34.5 gene, as well as a deletion of the UL39 gene coding for the large subunit of the ribonucleotide reductase. (See U.S. Pat. No. 5,585,096). G207 has been extensively studied in animal models and human phase I/II trials. Direct comparison between G207 and NV1020 revealed that NV1020 replicated more efficiently than G207 and exhibited higher oncolytic effectiveness at lower viral doses (McAuliffe et al., 2000; Cozzi et al., 2001; Bennett et al., 2002). Furthermore, G207 was noted to be not only attenuated for pathogenicity, but also for tumor cell killing capability (McAuliffe et al., 2000; Cozzi et al., 2001; Bennett et al., 2002).
HSV can be transmitted from cell-to-cell by causing limited amounts of virus-induced cell fusion, thus avoiding the extracellular environment. Specific mutations within viral glycoproteins are known to greatly enhance virus-induced cell fusion. Specifically, syncytial mutations that cause extensive virus-induced cell fusion can arise in at least two of the glycoprotein genes: the UL27gene, encoding glycoprotein B (gB) (Bzik et al., 1984a; Bzik et al., 1984b; Pellett et al., 1985; Manservigi, Spear, and Buchan, 1977), and the UL53 gene, coding for glycoprotein K (gK) (Bond and Person, 1984; Debroy, Pederson, and Person, 1985). Glycoprotein gK has been shown to function as a heterodimer with the UL20 viral protein and the UL20/gK heterodimer is necessary for virus-induced cell fusion (Foster et al., 2004; Melancon et al., 2007).
Breast cancer is the most common cancer among women, excluding cancers of the skin, accounting for nearly 1 in 3 cancers diagnosed in US women. In western countries breast cancer is the second leading cause of cancer death in women and is associated with high morbidity and mortality. A new and promising strategy for cancer therapy is the use of modified viruses that have been engineered to selectively replicate within cancer cells (oncolytic virotherapy). A number of viruses have been explored as tumor-selective replicating vectors, including adenovirus, herpes simplex virus type-1 (HSV-1), vaccinia virus, reovirus, Newcastle disease virus, vesicular stomatitis virus, measles virus, poliovirus and West Nile virus. Multiple murine tumor models have been used as preclinical settings for therapeutic purposes. The 4T1 mammary carcinoma model has several distinct advantages to be used as such model. It is regarded as a highly physiological, clinically-relevant mouse model that closely resembles stage 1V human breast cancer in its properties (Aslakson and Miller, 1992). 4T1 cells are considered to be very weakly immunogenic (relative antigenic strength is less than 0.01 with 9.9 being the most immunogenic), and they spontaneously metastasize to distal parts of the body (Aslakson and Miller, 1992; Pulaski and Ostrand-Rosenberg, 1998).
U.S. Pat. No. 5,328,688 discloses a recombinant herpes simplex virus vaccine based on the making the virus avirulent by prevention of expression of the γ1 34.5 gene.
International Publication No. WO 98/04726 discloses a herpes simplex virus strain that is disabled by inactivating both ICP34.5 and ICP27 genes for use as a gene delivery vector.
U.S. Patent Application Publication No. 2002/0019362 discloses treatment of cancers with a herpes simplex virus that has an alteration in the γ1 34.5 gene.
U.S. Pat. No. 6,846,670 discloses a genetically engineered herpes virus vector for treatment of cardiovascular disease that is modified by lacking a γ1 34.5 gene and operably comprises a heterologous nucleic acid.
U.S. Patent Application Publication No. 2007/0031383 discloses a recombinant herpes simplex virus expressing only a single γ1 34.5 gene and an expressible cytokine-encoding DNA.